A method for purifying nucleotides is provided, that includes preparing a solution comprising (a) 3'-blocked nucleotides, (b) 3'-OH nucleotides, (c) a polishing polymerase, and (d) a template. The polishing polymerase and the template are used to selectively polymerize the 3'-OH nucleotides and thus reduce a concentration in the solution of the 3 '-OH nucleotides relative to the 3'-blocked nucleotides.
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 21/04 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with deoxyribosyl as saccharide radical
C12P 19/34 - Polynucleotides, e.g. nucleic acids, oligoribonucleotides
2.
COMPOSITIONS AND METHODS FOR SEQUENCING BY SYNTHESIS
The present application relates to compositions and methods for sequencing by synthesis, where one or more palladium scavengers were used to improve sequencing metrics such phasing and prephasing values.
The present application relates to substituted dyes containing bis-boron fused heterocycles and their uses as fluorescent labels. These compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
A61K 47/54 - Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
A61K 31/196 - Carboxylic acids, e.g. valproic acid having an amino group the amino group being directly attached to a ring, e.g. anthranilic acid, mefenamic acid, diclofenac, chlorambucil
C09B 23/01 - Methine or polymethine dyes, e.g. cyanine dyes characterised by the methine chain
C09B 23/04 - Methine or polymethine dyes, e.g. cyanine dyes characterised by the methine chain containing an odd number of CH groups one CH group, e.g. cyanines, isocyanines, pseudocyanines
C09B 23/12 - Methine or polymethine dyes, e.g. cyanine dyes the polymethine chain being branched
An example of a flow cell includes a substrate and a pattern of two different silanes on at least a portion of a surface of the substrate. A first polymer is attached to a first of the two different silanes and a second polymer is attached to a second of the two different silanes. The first and second polymers respectively include a first functional group and a second functional group of a functional group pair, the functional group pair being selected from the group consisting of an activated ester functional group and an azide functional group, a tetrazine functional group and an activated ester functional group, and a tetrazine functional group and an azide functional group. A first primer set is grafted to the first polymer and a second primer set is grafted to the second polymer. The first and second primer sets are different.
Nucleic acid amplification techniques are disclosed. Embodiments include generating concatenated nucleic acids using rolling circle amplification of templates, e.g., starting from a cDNA of a full-length mRNA or from synthetic templates, and sequencing and/or detecting the concatenated nucleic acids. In some embodiments, the technology disclosed includes amplification reactions that include CRISPR-Cas interactions that generate primers as a result of the CRISPR-Cas interactions, whereby primers are in turn used as part of detectable amplification reactions. The disclosed amplification techniques may use synthetic oligonucleotides or primers.
The technology disclosed relates to efficiently determining which atoms in a protein are nearest to voxels in a grid. The atoms have three-dimensional (3D) atom coordinates, and the voxels have 3D voxel coordinates. The technology disclosed generates an atom-to-voxels mapping that maps, to each of the atoms, a containing voxel selected based on matching 3D atom coordinates of a particular atom of the protein to the 3D voxel coordinates in the grid. The technology disclosed generates a voxel-to-atoms mapping that maps, to each of the voxels, a subset of the atoms. The subset of the atoms mapped to a particular voxel in the grid includes those atoms in the protein that are mapped to the particular voxel by the atom-to-voxels mapping. The technology disclosed includes using the voxel-to-atoms mapping to determine, for each of the voxels, a nearest atom in the protein.
G16B 15/00 - ICT specially adapted for analysing two-dimensional or three-dimensional molecular structures, e.g. structural or functional relations or structure alignment
The present disclosure relates to a composition comprising a shell surrounding a core, wherein the core comprises one or more lyophilised microspheres. Also described herein is a method comprising providing one or more lyophilised microspheres; and coating the one or more lyophilised microspheres with a shell under conditions effective to encapsulate the one or more lyophilised microspheres. The present disclosure further relates to a system comprising one or more composition as described herein, and one or more lyophilised cake, wherein the one or more composition and the one or more lyophilised cake are combined under conditions effective to form a rehydration system. Also described herein is a method of controlling release of one or more encapsulated microspheres comprising providing a composition as described herein and mixing the composition with a rehydration solution under a first condition effective to control release of one or more lyophilised microspheres from the composition.
C12Q 1/6848 - Nucleic acid amplification reactions characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
The technology disclosed relates to determining pathogenicity of variants. In particular, the technology disclosed relates to generating amino acid-wise distance channels for a plurality of amino acids in a protein. Each of the amino acid-wise distance channels has voxel-wise distance values for voxels in a plurality of voxels. A tensor includes the amino acid-wise distance channels and at least an alternative allele of the protein expressed by a variant. A deep convolutional neural network determines a pathogenicity of the variant based at least in part on processing the tensor. The technology disclosed further augments the tensor with supplemental information like a reference allele of the protein, evolutionary conservation data about the protein, annotation data about the protein, and structure confidence data about the protein.
G16B 15/00 - ICT specially adapted for analysing two-dimensional or three-dimensional molecular structures, e.g. structural or functional relations or structure alignment
G16B 20/00 - ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations
A system includes at least a voxelizer, an alternative allele encoder, an evolutionary conservation encoder, and a convolutional neural network. The voxelizer accesses a three-dimensional structure of a reference amino acid sequence of a protein and fits a three-dimensional grid of voxels on atoms in the three-dimensional structure on an amino acid-basis to generate amino acid-wise distance channels. The alternative allele encoder encodes an alternative allele sequence to each voxel in the three-dimensional grid of voxels. The evolutionary conservation encoder encodes an evolutionary conservation sequence to each voxel in the three-dimensional grid of voxels. The convolutional neural network applies three-dimensional convolutions to a tensor that includes the amino acid-wise distance channels encoded with the alternative allele sequence and respective evolutionary conservation sequences and determines a pathogenicity of a variant nucleotide based at least in part on the tensor.
G16B 15/00 - ICT specially adapted for analysing two-dimensional or three-dimensional molecular structures, e.g. structural or functional relations or structure alignment
G16B 20/00 - ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations
METHODS OF PREPARING DIRECTIONAL TAGMENTATION SEQUENCING LIBRARIES USING TRANSPOSON-BASED TECHNOLOGY WITH UNIQUE MOLECULAR IDENTIFIERS FOR ERROR CORRECTION
Materials and methods for preparing nucleic acid libraries for next-generation sequencing are described herein. A variety of approaches are described relating to the use of unique molecular identifiers with transposon-based technology in the preparation of sequencing libraries. Also described herein are sequencing materials and methods for identifying and correcting amplification and sequencing errors.
A library sequencing technique with library quality control metrics is described. Sequence data using a sequencing primer that is complementary to a common adapter sequence in fragments of a nucleic acid sequencing library. The sequencing primer excludes a 3' terminal nucleotide of the common adapter sequence at a junction with a fragment insert. This exclusion avoids a mismatch region in any adapter dimers present in the sequencing library, and the sequence data includes adapter dimer sequence data, which is used to generate the quality control metrics.
A method for seeding and amplifying target nucleic acids derived from a sample in a cluster at a site on a surface of a substrate includes retaining at least a portion of the target nucleic acids in an inactive form that cannot seed to provide a relatively low concentration of active form target nucleic acids available for seeding. As the active form target nucleic acids seed on the surface of the substrate, they may be amplified. Because the concentration of active form target nucleic acids is low, the likelihood is low that a second active form target nucleic acid will seed at the same site within the same cluster before the first active form target nucleic acid is sufficiently amplified to dominate. Accordingly, the likelihood that the cluster will pass filters is increased relative to traditional seeding and amplification methods employing a higher concentration of active form target nucleic acids.
Examples provided herein are related to detecting methylcytosine and its derivatives using S- adenosyl-L-methionine analogs (xSAMs). Compositions and methods for performing such detection are disclosed. A target polynucleotide may include cytosine (C) and methylcytosine (mC). The method may include (a) protecting the C in the target polynucleotide from deamination; and (b) after step (a), deaminating the mC in the target polynucleotide to form thymine (T). Protecting the C from deamination may include adding a protective group to the 5 position of the C, e.g., using a methyltransferase enzyme that adds the first protective group from an xSAM.
Genomic library preparation using Cas-gRNA RNPs, and targeted epigenetic assays, are provided herein. Some compositions include, from a first species, substantially only single- stranded polynucleotides; from a second species, substantially only double-stranded polynucleotides; and amplification primers ligated to ends of the second double-stranded polynucleotides and substantially not ligated to any ends of the first double-stranded polynucleotides. Some compositions include first and second molecules of a target polynucleotide having a sequence, the first molecule having a first end at a first subsequence, the second molecule having a first end at a second subsequence, wherein the first subsequence only partially overlaps with the second subsequence. Some examples provide a composition that includes a target polynucleotide and a first fusion protein including a Cas- gRNA RNP coupled to a transposase having an amplification adapter coupled thereto. The Cas-gRNA RNP may be hybridized to a subsequence in the target polynucleotide.
The disclosure relates to methods, compositions, and kits for improving seeding efficiency of flow cells with polynucleotides, and applications thereof, including for sequencing.
In one example, an unsaturated cyclic dione is coupled to the substrate, and is reacted with an indole or indazole including a first functional group to form a first adduct coupling the first functional group to the substrate. In another example, an unsaturated cyclic dione is coupled to a substrate and reacted with a diene including a functional group to form an adduct coupling the functional group to the substrate. In another example, an indole or indazole is coupled to a substrate, and is reacted with an unsaturated cyclic dione including an oligonucleotide to form an adduct coupling the oligonucleotide to the substrate. In another example, a diene is coupled to a substrate, and is reacted with an unsaturated cyclic dione including an oligonucleotide to form an adduct coupling the oligonucleotide to the substrate.
Embodiments of the present disclosure relate to methods, kits and compositions for two- channel nuclei acid sequencing using blue and violet light excitation (e.g., lasers at 450-460 nm and 400-405 nm respectively). In particular, the nucleotides may be directly labeled with a blue dye, a violet dye, or both a blue dye and a violet dye. Alternatively, one or more nucleotides for incorporation may be unlabeled and affinity reagents containing a blue dye, a violet dye, or both a blue dye and a violet dye may be used to bind specifically to each type of nucleotides incorporated.
The present application relates to alkylpyridinium substituted coumarin dyes of formula (I) and their uses as fluorescent labels. For example, these dyes may be used to label nucleotides for nucleic acid sequencing applications. wherein R 1 is or and wherein R 1 is substituted with one or more C1-C6 alkyl.
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
C09B 62/00 - Reactive dyes, i.e. dyes which form covalent bonds with the substrates or which polymerise with themselves
C12Q 1/68 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
19.
METHODS, SYSTEMS AND COMPOSITIONS FOR NUCLEIC ACID SEQUENCING
The present disclosure relates to methods, systems, kits and compositions for nucleic acid sequencing applications. In particular, the method utilizes two imaging events with different excitation wavelengths and a single emission channel to collect the fluorescent signal patterns of different types of nucleotide conjugates to determine the identity of the incorporated nucleotide conjugates. The method described herein does not require a chemical treatment of the nucleotide conjugates in the incorporation mixture between the two imaging events.
The present application relates to long Stokes shift chromenoquinoline dyes and their uses as fluorescent labels. For example, these dyes may be used to label nucleotides for nucleic acid sequencing applications. The chromenoquinoline dyes have Formula (I).
C07D 457/14 - Heterocyclic compounds containing indolo [4, 3-f, g] quinoline ring systems, e.g. derivatives of ergoline, of the formula: , e.g. lysergic acid containing indolo [4, 3-f, g] quinoline ring systems condensed with carbocyclic rings or ring systems
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
C09B 57/00 - Other synthetic dyes of known constitution
G01N 33/50 - Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
An image sensor structure includes an image layer having an array of light detectors disposed therein. A device stack is disposed over the image layer. An array of light guides is disposed in the device stack. Each light guide is associated with a light detector. An array of nanowells is disposed over the device stack. Each nanowell is associated with a first light guide of the array of light guides. A first primer set is disposed throughout a first well region of each nanowell. A second primer set is disposed throughout a second well region of each nanowell. The second well region is adjacent the first well region. The first and second primer sets are operable to attach a forward strand cluster of forward polynucleotide strands in the first well region and a reverse strand cluster of reverse polynucleotide strands in the second well region.
An example of an incorporation mix includes a liquid carrier, a complex, and a labeled nucleotide. The complex includes a polymerase and a plasmonic nanostructure linked to the polymerase. The labeled nucleotide includes a nucleotide, a 3' OH blocking group attached to a sugar of the nucleotide, and a dye label attached to a base of the nucleotide.
An example of a functionalized plasmonic nanostructure includes a plasmonic nanostructure core; a polymeric hydrogel attached to the plasmonic nanostructure core, the polymeric hydrogel having a thickness ranging from about 10 nm to about 200 nm; and a plurality of primers attached to side chains or arms of the polymeric hydrogel, wherein at least some of the plurality of primers are attached to the polymeric hydrogel at different distances from the plasmonic nanostructure core.
In an example method, a hydrogel is applied to a surface of a substrate and primers are grafted to the applied hydrogel. Before or after the primers are grafted, plasmonic nanostructures are introduced to the applied hydrogel. This substrate can make up one surface of a flow cell. When the flow cell is used in a sequencing operation, the plasmonic nanostructures can enhance fluorescent signals that are generated.
A method is used to generate an analysis image of a moving sample based on one or more exposures. An illumination source illuminates a field of view of a camera for one or more pulses while the sample moves through the field of view. The distance moved by the sample during each of these one or more pulses may be less than the size of one pixel in an image captured by the camera.
Described herein is a polynucleotide for use as a sequencing template comprising multiple inserts. Also described herein are method of generating and using these polynucleotides and methods of use of such templates, including analysis of contiguity information. Further, sequencing templates comprising an insert sequence and a copy of the insert sequence can be used to correct for random errors generated during sequencing or amplification or to identify nucleobase damage or other mutation that leads to non-canonical base pairing in a double-stranded nucleic acid. Methods of performing methylation analysis are also described herein.
A composition for amplifying a polynucleotide is provided that includes a substrate comprising a first region and a second region. A first plurality of capture primers is coupled to the first region of the substrate. A second plurality of capture primers is coupled to the second region of the substrate. The capture primers of the second plurality of capture primers are longer than the capture primers of the first plurality of capture primers. A first plurality of orthogonal capture primers are coupled to the first region of the substrate. A second plurality of orthogonal capture primers are coupled to the second region of the substrate. The orthogonal capture primers of the second plurality of orthogonal capture primers are shorter than the orthogonal capture primers of the first plurality of orthogonal capture primers.
Disclosed herein is a method for enriching a sequencing library comprising double-stranded nucleic acid fragments comprising preparing a library of double-stranded fragments having one or more adaptors at ends of the double-stranded fragment; denaturing the double-stranded fragments to form single-stranded fragments; and hybridizing an extension primer that binds to a target sequence of at least one insert in the library of double-stranded fragments and that does not bind to non-target sequences. In an embodiment, the adaptor is a hairpin adaptor, and extension from the extension primer using a polymerase with 5' to 3' exonuclease activity removes all or part of a sequence of the hairpin adaptor that is at least partially complementary to the amplification primer sequence.
A variety of different types of targeted transposome complexes are described herein that may be used to mediate sequence-specific targeted transposition of nucleic acids. Also described herein is a method of characterizing desired samples in a mixed pool of samples comprising both desired samples and unwanted samples comprising, to produce sequencing data from double-stranded nucleic acid, initially sequencing a library comprising a plurality of nucleic acid samples from a mixed pool, wherein each nucleic acid library comprises nucleic acids from a single sample and a unique sample barcode to distinguish the nucleic acids from the single sample from the nucleic acids from other samples in the library; analyzing the sequencing data and identifying unique sample barcodes associated with sequencing data from desired samples; performing a selection step on the library comprising enriching nucleic acid samples from desired samples and/or depleting nucleic acid samples from unwanted samples; and resequencing the nucleic acid library.
This application describes methods of preparing an immobilized library of tagged RNA fragments. Also described herein are a number of methods of preparing DNA and RNA sequencing libraries from a single sample. These methods can include library preparation from single cells.
The present application relates to substituted coumarin derivatives and their uses as fluorescent labels. These compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
C07D 403/04 - Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 405/04 - Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 405/14 - Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing three or more hetero rings
C07D 407/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 409/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 417/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
C09B 62/465 - Reactive dyes, i.e. dyes which form covalent bonds with the substrates or which polymerise with themselves with the reactive group not directly attached to a heterocyclic ring the reactive group being an acryloyl group, a quaternised or non-quaternised aminoalkyl carbonyl group, or a (—N)n—CO—A—O—X or (—N)n—CO—A—Hal group, wherein A is an alkylene or alkylidene group, X is hydrogen or an acyl radical of an organic or inorg
Provided is a method, including stretching a polynucleotide over a substrate including a plurality of equally spaced cleavage regions including a plurality of transposases, cleaving the polynucleotide with two or more of the plurality of transposases to form a plurality of polynucleotide fragments, and separating, within the plurality of polynucleotide fragments, a population of longer polynucleotide fragments from a population of shorter polynucleotide fragments. Also provided is a method including stretching a polynucleotide over a substrate including a plurality of equally spaced cleavage regions including a plurality of transposases, cleaving the polynucleotide with two or more of the plurality of transposases to form a plurality of polynucleotide fragments, and separating, within the plurality of polynucleotide fragments, a population of longer polynucleotide fragments from a population of shorter polynucleotide fragments.
The present invention concerns a nucleoside or nucleotide comprising a nucleobase attached to a detectable label via a cleavable linker, wherein the nucleoside or nucleotide comprises a ribose or 2' deoxyribose moiety and a 3'-OH blocking group, and wherein the cleavable linker comprises a moiety of the structure (I) wherein each of X and Y is independently O or S; and each of R1a, R2b, R2, R3a and R3b is independently H, halogen, unsubstituted or substituted C1-C6 alkyl, or C1-C6 haloalkyl. Also provided herein are methods to prepare such nucleotide and nucleoside molecules, and the uses of fully functionalized nucleotides containing the 3' acetal blocking group for sequencing applications.
C12Q 1/6874 - Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation [SBH]
C07H 19/10 - Pyrimidine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 21/02 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with ribosyl as saccharide radical
C07H 21/04 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with deoxyribosyl as saccharide radical
34.
METHODS FOR INCREASING YIELD OF SEQUENCING LIBRARIES
The present disclosure is concerned with compositions and methods for preparing a sequencing library. In one embodiment, methods include producing a library of target nucleic acids having the same adapter at each end and then switching the identity of one adapter to result in target nucleic acids flanked by distinct adapters.
An example of a flow cell includes a substrate and a cured, patterned resin on the substrate. The cured, patterned resin has nano-depressions separated by interstitial regions. Each nano-depression has a largest opening dimension ranging from about 10 nm to about 1000 nm. The cured, patterned resin also includes an interpenetrating polymer network. The interpenetrating polymer network of the cured, patterned resin includes an epoxy-based polymer and a (meth)acryloyl-based polymer.
B32B 3/30 - Layered products essentially comprising a layer with external or internal discontinuities or unevennesses, or a layer of non-planar form; Layered products essentially having particular features of form characterised by a layer with cavities or internal voids characterised by a layer formed with recesses or projections, e.g. grooved, ribbed
B82Y 15/00 - Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors
B29C 64/124 - Processes of additive manufacturing using only liquids or viscous materials, e.g. depositing a continuous bead of viscous material using layers of liquid which are selectively solidified
B32B 37/15 - Methods or apparatus for laminating, e.g. by curing or by ultrasonic bonding characterised by the properties of the layers with at least one layer being manufactured and immediately laminated before reaching its stable state, e.g. in which a layer is extruded and laminated while in semi-molten state
C08J 3/24 - Crosslinking, e.g. vulcanising, of macromolecules
C08L 33/00 - Compositions of homopolymers or copolymers of compounds having one or more unsaturated aliphatic radicals, each having only one carbon-to-carbon double bond, and only one being terminated by only one carboxyl radical, or of salts, anhydrides, esters,; Compositions of derivatives of such polymers
C08L 63/00 - Compositions of epoxy resins; Compositions of derivatives of epoxy resins
C12Q 1/68 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
An example of a kit includes a flow cell and a genotyping probe fluid. The flow cell includes a substrate, and first and second capture primers attached to the substrate. The genotyping probe fluid includes a liquid carrier, and a genotyping oligonucleotide in the liquid carrier. The genotyping oligonucleotide includes a first primer sequence; a probe sequence that is representative of a target genotyping locus; a restriction endonuclease site; and a second primer sequence that is at least partially complementary to the second capture primer.
Provided is a nanoparticle including a scaffold, a single template site for bonding a template polynucleotide to the scaffold, and a plurality of accessory sites for bonding accessory oligonucleotides to the scaffold, wherein the scaffold is selected from an asymmetrical acrylamide polymer one or a dendrimer including lysyl constitutional repeating units, the single template site for bonding a template polynucleotide to the scaffold is selected from a covalent template bonding site and a noncovalent template bonding site and the plurality of accessory sites for bonding accessory oligonucleotides to the scaffold are selected from covalent accessory oligonucleotide bonding sites and noncovalent accessory oligonucleotide bonding sites. Also provided are methods of using the nanoparticle.
C40B 50/14 - Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
38.
NANOPARTICLE WITH SINGLE SITE FOR TEMPLATE POLYNUCLEOTIDE ATTACHMENT
Provided is a nanoparticle including a scaffold, a single template site for bonding a template polynucleotide to the scaffold, and a plurality of accessory sites for bonding accessory oligonucleotides to the scaffold, wherein the scaffold is selected from one or more scaffold DNA molecules and one or more scaffold polypeptides, the single template site for bonding a template polynucleotide to the scaffold is selected from a covalent template bonding site and a noncovalent template bonding site and the plurality of accessory sites for bonding accessory oligonucleotides to the scaffold are selected from covalent accessory oligonucleotide bonding sites and noncovalent accessory oligonucleotide bonding sites. Also provided are methods of using the nanoparticle, in particular in sequencing by synthesis.
C40B 50/14 - Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
An example of a kit includes a flow cell and a cleavage mix. An example flow cell includes a substrate; a catalytic polymeric hydrogel on the substrate, the catalytic polymeric hydrogel including a deblocking catalyst; and an amplification primer attached to the catalytic polymeric hydrogel. The deblocking catalyst accelerates cleavage of a blocking group of a 3' OH blocked nucleotide introduced to the flow cell and incorporated into a template strand attached to the amplification primer. An example of the cleavage mix includes a component to initiate cleavage of the blocking group.
In an example method, a series of time-based clustering images is generated for a plurality of library fragments from a genome sample. Each time-based clustering image in the series is sequentially generated. To generate each time-based clustering image in the series: i) a respective sample is introduced to a flow cell, the respective sample including contiguity preserved library fragments of the plurality of library fragments, wherein the contiguity preserved library fragments are attached to a solid support or are attached to each other; ii) the contiguity preserved library fragments are released from the solid support or from each other; iii) the contiguity preserved library fragments are amplified to generate a plurality of respective template strands; iv) the respective template strands are stained; and v) the respective template strands are imaged.
A hydrogel includes a dendritic core with 2 to 30 arms, and first and second acrylamide monomers incorporated into each arm. The first acrylamide monomer is: (I), wherein R1 and R2 are independently selected from an alkyl, an alkylamino, an alkylamido, an alkylthio, an aryl, a glycol, and optionally substituted variants thereof; and the second acrylamide monomer is: (II), wherein R3 and R4 are independently hydrogen or an alkyl; L is a linker including a linear chain of 2 to 20 atoms selected from carbon, oxygen, and nitrogen and optional substituents on the carbon and any nitrogen atoms; A is an N substituted amide: (III), where R5 is hydrogen or an alkyl; E is a linear chain of 1 to 4 atom(s) selected from carbon, oxygen and nitrogen, and optional substituents on the carbon and any nitrogen atoms; and Z is an optional nitrogen containing heterocycle.
Embodiments of the present disclosure relate to cyclooctatetraene (COT) containing dyes and their uses as fluorescent labels. The fluorescent compounds comprise photo-protecting cyclooctatetraene moiety of formula (I). Also provided are compositions containing cyclooctatetraene (COT). The dyes and compositions may be used in various biological applications, such as nucleic acid sequencing.
C07H 21/04 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with deoxyribosyl as saccharide radical
C09B 1/00 - Dyes with an anthracene nucleus not condensed with any other ring
A method of generating an asymmetric closed-ended double stranded nucleic acid template from a double stranded nucleic acid template having free 5' and 3' ends by use of hairpin or dumbbell adaptors, and sequencing therefrom.
A method for detecting different analytes includes mixing different analytes with sensing probes, wherein at least some of the sensing probes are specific to respective ones of the analytes. The analytes respectively are captured by the sensing probes that are specific to those analytes. Fluorophores respectively are coupled to sensing probes that captured respective analytes. The sensing probes are mixed with beads, wherein the beads are specific to respective ones of the sensing probes, and wherein the beads include different codes identifying the analytes to which those sensing probes are specific. The sensing probes respectively are coupled to beads that are specific to those sensing probes. The beads are identified that are coupled to the sensing probes that captured analytes using at least fluorescence from the fluorophores coupled to those sensing probes. The analytes that are captured are identified.
Presented are methods and compositions for using immobilized CRISPR/Cas9 enzymes for generating an immobilized library of randomly fragmented, double-stranded target nucleic acid fragments on a surface. The methods are useful for generating nucleic acid fragments for use in a variety of processes, including massively parallel nucleic acid sequencing.
C12N 11/00 - Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
C12N 15/10 - Processes for the isolation, preparation or purification of DNA or RNA
C12N 15/11 - DNA or RNA fragments; Modified forms thereof
C12Q 1/68 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
C40B 40/06 - Libraries containing nucleotides or polynucleotides, or derivatives thereof
C40B 50/14 - Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
46.
NUCLEIC ACID LIBRARY PREPARATION USING ELECTROPHORESIS
Described herein are methods and systems for performing chemical or enzymatic reactions using electrophoresis. Devices, systems, and methods for preparing a library of tagged nucleic acid fragments from a target double-stranded nucleic acid using electrophoresis are also provided. Application of one or more electric fields causes molecules to migrate through the electrophoresis gel matrix.
For example, a flowcell includes: a nanowell layer having a first set of nanowells and a second set of nanowells to receive a sample; a first linear waveguide associated with the first set of nanowells, and a second linear waveguide associated with the second set of nanowells; and a first grating for the first linear waveguide, and a second grating for the second linear waveguide, the first and second gratings providing differential coupling of first light and second light.
Embodiments of the present disclosure relate to nucleotides labeled with photoswitchable compounds. Also provided herein are methods and kits of using these labeled nucleotides for sequencing applications.
C12Q 1/6874 - Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation [SBH]
C07H 19/00 - Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro derivatives thereof
In a first aspect, a method includes:providing a sample, the sample including a first nucleotide and a second nucleotide;contacting the sample with a first fluorescent dye and a second fluorescent dye, the first fluorescent dye emitting first emitted light within a first wavelength band responsive to a first excitation illumination light, the second fluorescent dye emitting second emitted light within a second wavelength band responsive to a second excitation illumination light; simultaneously collecting, using one or more image detectors, multiplexed fluorescent light comprising the first emitted light and the second emitted light, the first emitted light being a first color channel corresponding to the first wavelength band and the second emitted light being a second color channel corresponding to the second wavelength band; and identifying the first nucleotide based on the first wavelength band of the first color channel and the second nucleotide based on the second wavelength band of the second color channel.
The present application relates to tertiary amine substituted coumarin derivatives of formula (I) and their uses as fluorescent labels. These compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
C07D 405/04 - Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 413/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 417/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07H 19/10 - Pyrimidine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
The present application relates to exocyclic amine-substituted coumarin derivatives and their uses as fluorescent labels. These compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
C07D 417/14 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group containing three or more hetero rings
C07D 405/14 - Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing three or more hetero rings
C07D 413/14 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing three or more hetero rings
C07H 19/10 - Pyrimidine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
A flow cell includes a support and a heteropolymer attached to the support. The heteropolymer includes an acrylamide monomer including an attachment group to react with a functional group attached to a primer, and a monomer including a stimuli-responsive functional group. The monomer including the stimuli-responsive functional group may be pH-responsive, temperature-responsive, saccharide-responsive, nucleophile-responsive, and/or salt-responsive.
C12Q 1/6874 - Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation [SBH]
B01L 3/00 - Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
C08F 220/20 - Esters of polyhydric alcohols or phenols
C08F 220/60 - Amides containing nitrogen in addition to the carbonamido nitrogen
C08F 230/06 - Copolymers of compounds having one or more unsaturated aliphatic radicals, each having only one carbon-to-carbon double bond, and containing phosphorus, selenium, tellurium, or a metal containing a metal containing boron
The present disclosure relates to methods, compositions, and kits for generating a library of tagged nucleic acid fragments without using PCR amplification, including methods and compositions for fragmenting and tagging nucleic acids (e.g., DNA) using transposome complexes immobilized on solid support.
Embodiments of the present disclosure relate to nucleotide and nucleoside molecules with acetal or thiocarbamate 3'-OH blocking groups. Also provided herein are methods to prepare such nucleotide and nucleoside molecules, and the uses of fully functionalized nucleotides containing the 3'-OH blocking group for sequencing applications.
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
C12Q 1/68 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
A polynucleotide sequencing method comprises (i) removing a label and a blocking moiety from a blocked, labeled nucleotide incorporated into a copy polynucleotide strand that is complementary to at least a portion of a template polynucleotide strand; and (ii) washing the removed label and blocking moiety away from the copy strand with a wash solution comprising a first buffer comprising a scavenger compound. Removing the label and blocking moieties may comprise chemically removing the moieties. The first buffer may also comprise an antioxidant and may be used in a scanning buffer used during a nucleotide detection step.
Polynucleotide sequencing methods employ a sequencing oligonucleotide that hybridizes to a free 3' end potion of a template polynucleotide strand with greater affinity than a surface oligonucleotide. Such sequencing oligonucleotides may be used as a primer to determine the sequence of an index sequence by extending the sequencing oligonucleotide using the template strand as a template. Sequencing processes that employ such sequencing oligonucleotides provide a sufficiently intense signal to determine to the sequence of the index sequence.
The present disclosure is concerned with compositions and methods for the paired-end sequencing of target nucleic acids, and more particularly to obtaining nucleotide sequence information from two separate regions of target nucleic acids using amplification sites having a single type of surface primer.
Described herein are heterocyclic azide-containing monomer units, copolymers comprising such heterocyclic azide-containing monomer units, substrate- bound copolymers, and oligonucleotide-bound copolymers, methods for making such copolymers and reacting them with a substrate and/or oligonucleotide, and methods of using such copolymers for immobilization of oligonucleotides to a substrate, for example for use in DNA sequencing or other diagnostic applications.
C07D 207/16 - Carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
C07D 213/75 - Amino or imino radicals, acylated by carboxylic or carbonic acids, or by sulfur or nitrogen analogues thereof, e.g. carbamates
C07D 401/06 - Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms
C08F 8/30 - Introducing nitrogen atoms or nitrogen-containing groups
C08F 293/00 - Macromolecular compounds obtained by polymerisation on to a macromolecule having groups capable of inducing the formation of new polymer chains bound exclusively at one or both ends of the starting macromolecule
59.
DECREASING PHASING WITH UNLABELED NUCLEOTIDES DURING SEQUENCING
Polynucleotide sequencing methods include incubating unlabeled nucleotides with a cluster of template polynucleotide strands having the same sequence when the identity of the previously added labeled nucleotide is being detected. The detection step provides time for the addition of the unlabeled nucleotides to be incorporated into the copy strands in which the previously added labeled nucleotide did not get incorporated. Thus, at the end of the detection step, all or most of the copy strands will be in phase and ready to incorporate the appropriate labeled nucleotide in the subsequence incorporate step.
The present disclosure is concerned with compositions and methods for reducing the steps used in the generation of monoclonal clusters by combining the enzymes used for linearization and removal of unused surface primers.
Presented herein are altered polymerase enzymes for improved incorporation of nucleotides and nucleotide analogues, in particular altered polymerases that maintain high fidelity under reduced incorporation times, as well as methods and kits using the same.
Disclosed are methods for determining a genetic origin of fetal cellular DNA obtained from a pregnant female who is carrying a fetus in a current pregnancy. Methods are also disclosed for using the fetal cellular DNA and fetal cell-free DNA (cfDNA) to determine fetal genetic conditions such as copy number variations. The methods disclosed uses a probabilistic model to determine fetal cellular DNA origin based on alleles observed at informative genetic marker of the fetal cellular DNA. Systems and computer program products for performing the methods are also disclosed.
This disclosure describes a hybridization buffer including a crowding agent, a method that includes using the hybridization buffer, and a kit including the hybridization buffer. This disclosure also describes blockers for use in hybrid capture methods, methods of using those blockers, and a kit including those blockers. Additionally, this disclosure describes a method of hybrid capture that does not include amplifying the library members using PCR prior to sequencing the library members.
In an example, a flow cell includes a substrate, a selectively removable porous molecular network on the substrate and defining exposed substrate regions, and sequencing surface chemistry on at least some of the exposed regions. The sequencing surface chemistry is selected from the group consisting of i) an activated pad, a polymer layer attached to the activated pad, and a primer attached to the polymer layer; or ii) a nanostructure and an enzyme attached to the nanostructure.
B01J 19/00 - Chemical, physical or physico-chemical processes in general; Their relevant apparatus
C12Q 1/6809 - Methods for determination or identification of nucleic acids involving differential detection
C12Q 1/00 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
C23C 28/00 - Coating for obtaining at least two superposed coatings either by methods not provided for in a single one of main groups , or by combinations of methods provided for in subclasses and
65.
RESIN COMPOSITION AND FLOW CELLS INCORPORATING THE SAME
An example of a resin composition includes a free radical curable resin matrix including an acrylate and a siloxane, and a free radical photoinitiator. When cured, the resin composition has low or no autofluorescence when exposed to blue excitation wavelengths ranging from about 380 nm to about 480 nm or green excitation wavelengths ranging from about 510 nm to about 560 nm.
An example of a resin composition includes an epoxy resin matrix, a free radical photoinitiator selected from the group consisting of 2-ethyl-9, 10-dimethoxyanthracene, 2,2-dimethoxy-2-phenylacetophenone, 2-ethoxy-2-phenylacetophenone, and a phosphine oxide, and a photoacid generator. When cured, the resin composition has low or no autofluorescence when exposed to blue excitation wavelengths ranging from about 380 nm to about 480 nm or green excitation wavelengths ranging from about 510 nm to about 560 nm.
G03F 7/033 - Non-macromolecular photopolymerisable compounds having carbon-to-carbon double bonds, e.g. ethylenic compounds with binders the binders being polymers obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. vinyl polymers
An example of a flow cell includes a substrate; a first primer set attached to a first region on the substrate, the first primer set including an un-cleavable first primer and a cleavable second primer; and a second primer set attached to a second region on the substrate, the second primer set including a cleavable first primer and an un- cleavable second primer.
Embodiments of the present disclosure relate to methods of preparation of templates for polynucleotide sequencing. In particular, the disclosure relates to linearization of clustered polynucleotides in preparation for sequencing by cleavage of one or more first strands of double-stranded polynucleotides immobilized on a solid support by a transition metal complex, for example, a palladium complex or a nickel complex. Further disclosure relate to linearization of clustered polynucleotides by cleaving one or more second strands of double double-stranded polynucleotides immobilized on a solid support comprising azobenzene linker by Na2S2O4. Nucleotides and oligonucleotides comprising a 3 ' phosphate moiety blocking group, and methods of removing the same using a fluoride reagent are also disclosed.
C07H 21/04 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with deoxyribosyl as saccharide radical
C12Q 1/6806 - Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
C12Q 1/6874 - Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation [SBH]
A method of sequencing a plurality of polynucleotides includes: attaching a single DNA template molecule to each of a plurality of attachment elements on a sample container, wherein the average distance between adjacent elements is less than Abbe's limit; applying a stochastic photo-switching chemistry to all of the molecules at the same time to cause the attached molecules to fluoresce in on and off events in up to four different colors by stochastic photo-switching; and imaging the on and off events in a color channel for each color in real-time as the on and off events are occurring for the attached molecules.
Techniques are described for reducing the number of angles needed in structured illumination imaging of biological samples through the use of patterned flowcells, where nanowells of the patterned flowcells are arranged in, e.g., a square array, or an asymmetrical array. Accordingly, the number of images needed to resolve details of the biological samples is reduced. Techniques are also described for combining structured illumination imaging with line scanning using the patterned flowcells.
An apparatus for directing fluid into and out of a fluidic device includes two or more fluid prime channels connected to a fluid inlet of the fluidic device, a flow control valve for each fluid prime channel to control flow between the fluid prime channel and the fluid inlet, one or more outlet channels connected to a fluid outlet of the fluidic device, and a flow control valve for each outlet channel to control flow between the fluid outlet and the associated outlet channel. An apparatus for delivering fluids to a fluid inlet includes a plate that is rotatable about an axis of rotation and a plurality of fluid compartments disposed on the plate, each compartment having a fluid exit port disposed at a common radial distance from the axis of rotation and positioned to align with the fluid inlet as the plate rotates about the axis of rotation.
G01N 35/08 - Automatic analysis not limited to methods or materials provided for in any single one of groups ; Handling materials therefor using a stream of discrete samples flowing along a tube system, e.g. flow injection analysis
G01N 35/02 - Automatic analysis not limited to methods or materials provided for in any single one of groups ; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations
G01N 37/00 - INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES - Details not covered by any other group of this subclass
The technology disclosed directly operates on sequencing data and derives its own feature filters. It processes a plurality of aligned reads that span a target base position. It combines elegant encoding of the reads with a lightweight analysis to produce good recall and precision using lightweight hardware. For instance, one million training examples of target base variant sites with 50 to 100 reads each can be trained on a single GPU card in less than 10 hours with good recall and precision. A single GPU card is desirable because it a computer with a single GPU is inexpensive, almost universally within reach for users looking at genetic data. It is readily available on could-based platforms.
G16B 20/20 - Allele or variant detection, e.g. single nucleotide polymorphism [SNP] detection
G16B 40/00 - ICT specially adapted for biostatistics; ICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
In an example of the method, a functionalized coating layer is applied in depressions of a patterned flow cell substrate. The depressions are separated by interstitial regions. A primer is grafted to the functionalized coating layer to form a grafted functionalized coating layer in the depressions. A hydrogel is applied on at least the grafted functionalized coating layer.
The present application relates to secondary amine-substituted coumarin compounds and their uses as fluorescent labels. The compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications. In one embodiment, the compound is a compound of Formula (I) or a salt thereof:
C07D 405/04 - Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 413/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 417/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
The assembly includes a docking console and a manifold. The docking console includes a cartridge support surface having a first end and a second end. The manifold has one or more wells defined therein The docking console further includes a manifold retention bracket to releasably hold the manifold against a fluid cartridge supported on the cartridge support surface at an interface position such that the one or more wells are in fluid communication with the fluid cartridge and a biased seal bar to press the fluid cartridge against the manifold held by the manifold retention bracket. A hydrophilic porous frit disposed within at least one of the wells and is to permit liquid to flow through the outlet aperture but prevent gas from passing through the outlet aperture.
The present disclosure relates to new nucleotide and oligonucleotide compounds and their use in nucleic acid sequencing applications. The compounds have formula (I) wherein B is a nucleoside base; and Fl is a fluorophore attached through an optional linker. More specifically the compounds have formulas (c) or (t) wherein p is a triphosphate group; and Fl is a fluorophore attached through an optional linker.
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
An example of a method includes modifying an exposed surface of a substrate to incorporate a first chemical group; reacting the first chemical group with a first reactive group of a functionalized polymer molecule to form a functionalized polymer coating layer covalently bound to the exposed surface of the substrate; grafting a primer to the functionalized polymer coating layer by reacting the primer with a second reactive group of the functionalized polymer coating layer; and forming a water-soluble protective coating on the primer and the functionalized polymer coating layer. Examples of flow cells incorporating examples of the water-soluble protective coating are also disclosed herein.
The present invention is concerned with compositions and methods for improving the rate of correct sample identification in indexed nucleic acid library preparations for multiplex next generation sequencing by modifying or blocking 5' and 3' ends of pooled indexed polynucleotides from multiple samples, with an optional exonuclease treatment, prior to amplification and sequencing.
The present invention is concerned with compositions and methods for improving the rate of correct sample identification in indexed nucleic acid library preparations for multiplex next generation sequencing by exonuclease treatment after protective adapters are ligated to target polynucleotides to degrade unincorporated adapters prior to amplification and sequencing.
The present invention is concerned with compositions and methods for improving the rate of correct sample identification in indexed nucleic acid library preparations for multiplex next generation sequencing by exonuclease treatment and optionally blocking the 3' ends of pooled indexed polynucleotides from multiple samples prior to amplification and sequencing.
The present invention is concerned with compositions and methods for improving the rate of correct sample identification in indexed nucleic acid library preparations for multiplex next generation sequencing by blocking the 3' ends of pooled indexed polynucleotides from multiple samples prior to amplification and sequencing.
Disclosed is a system for determining the nucleotide sequence of polynucleotides. The system can comprise a light source, such as a laser or a LED, configured to generate light at a predetermined wavelength. A detector of the system can detect fluorescent emissions at a first wavelength and a second wavelength. A processor of the system identify the nucleotide as a first type if no fluorescent emission is detected by the at least one detector; identify the nucleotide as a second type if a fluorescent emission at the first wavelength of light is detected by the at least one detector; identify the nucleotide as a third type if a fluorescent emission at the second wavelength of light is detected by the at least one detector; and identify the nucleotide as a fourth type if fluorescent emissions at the first wavelength and the second wavelength of light are detected by the at least one detector.
The present disclosure relates to methods, compositions, and kits for treating target nucleic acids, including methods and compositions for fragmenting and tagging nucleic acid (e.g., DNA) using transposome complexes bound to a solid support.
An example method includes reacting a first solution and a different, second solution on a flow cell by flowing the first solution over amplification sites on the flow cell and subsequently flowing the second solution over the amplification sites. The first solution includes target nucleic acids and a first reagent mixture that comprises nucleoside triphosphates and replication enzymes. The target nucleic acids in the first solution transport to and bind to the amplification sites at a transport rate. The first reagent mixture amplifies the target nucleic acids that are bound to the amplification sites to produce clonal populations of amplicons originating from corresponding target nucleic acids. The amplicons are produced at an amplification rate that exceeds the transport rate. The second solution includes a second reagent mixture and lacks the target nucleic acids. The second solution is to increase a number of the amplicons at the amplification sites.
An imprinting apparatus includes a silicon master having a plurality of nanofeatures defined therein. An anti-stick layer coats the silicon master, the anti-stick layer including a molecule having a cyclosiloxane with at least one silane functional group. A method includes forming a master template by: depositing a formulation on a silicon master including a plurality of nanofeatures defined therein, the formulation including a solvent and a molecule having a cyclosiloxane with at least one silane functional group; and curing the formulation, thereby forming an anti-stick layer on the silicon master, the anti-stick layer including the molecule. The method further includes depositing a silicon-based working stamp material on the anti-stick layer of the master template; curing the silicon-based working stamp material to form a working stamp including a negative replica of the plurality of nanofeatures; and releasing the working stamp from the master template.
G03F 7/00 - Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printed surfaces; Materials therefor, e.g. comprising photoresists; Apparatus specially adapted therefor
An example of an array includes a support, a cross-linked epoxy polyhedral oligomeric silsesquioxane (POSS) resin film on a surface of the support, and a patterned hydrophobic polymer layer on the cross-linked epoxy POSS resin film. The patterned hydrophobic polymer layer defines exposed discrete areas of the cross-linked epoxy POSS resin film, and a polymer coating is attached to the exposed discrete areas. Another example of an array includes a support, a modified epoxy POSS resin film on a surface of the support, and a patterned hydrophobic polymer layer on the modified epoxy POSS resin film. The modified epoxy POSS resin film includes a polymer growth initiation site, and the patterned hydrophobic polymer layer defines exposed discrete areas of the modified epoxy POSS resin film. A polymer brush is attached to the polymer growth initiation site in the exposed discrete areas.
A flow cell package includes first and second surface-modified patterned wafers and a spacer layer. The first surface-modified patterned wafer includes first depressions separated by first interstitial regions, a first functionalized molecule bound to a first silane or silane derivative in at least some of the first depressions, and a first primer grafted to the first functionalized molecule in the at least some of the first depressions. The second surface-modified patterned wafer includes second depressions separated by second interstitial regions, a second functionalized molecule bound to a second silane or silane derivative in at least some of the second depressions, and a second primer grafted to the second functionalized molecule in the at least some of the second depressions. The spacer layer bonds at least some first interstitial regions to at least some second interstitial regions, and at least partially defines respective fluidic chambers of the flow cell package.
H01L 21/3213 - Physical or chemical etching of the layers, e.g. to produce a patterned layer from a pre-deposited extensive layer
C07C 247/12 - Compounds containing azido groups with azido groups bound to acyclic carbon atoms of a carbon skeleton being further substituted by carboxyl groups
C12Q 1/68 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
The present application relates to new coumarin compounds and their uses as fluorescent labels. The compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
Disclosed herein are systems and methods for spinal muscular atrophy (SMA) diagnosis from whole genome sequencing data. In one embodiment, a method comprises aligning whole genome sequencing (WGS) reads of a subject's sample to a modified reference sequence such as a modified reference genome sequence. After counting the reads supporting quasi-alleles at select positions of the reference sequence, the method can adjust for coverage and determine a number of functional SMNl gene copies. The method can determine affected or carrier status of the subject based on the copy number of functional SMNl gene copies.
C12Q 1/6883 - Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
The present application relates to fluorescent dyes of formula (l) and their uses as fluorescent labels. The compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
C07D 417/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
Example super-resolution microscopy systems are described herein that are configured for relatively high throughput. The disclosed microscopy systems can be to generate an array of sub-diffraction activated areas for imaging. The microscopy systems can be to utilize imaging techniques that employ time delay integration to build up super- resolution images over time. The disclosed microscopy systems can utilize long-lived fluorophores in conjunction with wide field and patterned illumination to generate super- resolution images of a sample with relatively high throughput.
Disclosed herein are compositions and fluidic devices that include a filler fluid having a siloxane block co-polymer solubilized in the filler fluid. Also disclosed herein are related kits and methods for using the fluidic devices for various uses, such as the polymerase chain reaction or preparations for sequencing reactions.
The present disclosure relates to new polymethine compounds and their use as fluorescent markers. In particular the compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications. In one embodiment, a compound of the formula (I) or mesomeric forms thereof is provided: (see formula I)
C07D 403/06 - Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms
C07D 209/08 - Indoles; Hydrogenated indoles with only hydrogen atoms or radicals containing only hydrogen and carbon atoms, directly attached to carbon atoms of the hetero ring
C09B 23/02 - Methine or polymethine dyes, e.g. cyanine dyes characterised by the methine chain containing an odd number of CH groups
C09B 23/06 - Methine or polymethine dyes, e.g. cyanine dyes characterised by the methine chain containing an odd number of CH groups three CH groups, e.g. carbocyanines
G01N 33/00 - Investigating or analysing materials by specific methods not covered by groups
G01N 33/533 - Production of labelled immunochemicals with fluorescent label
94.
ENHANCED UTILIZATION OF SURFACE PRIMERS IN CLUSTERS
Presented herein are methods and compositions for enhancing utilization of surface primers during the surface amplification process. The methods are useful for surface amplification at improved densities. The methods and compositions provided herein enable creation of clusters which are brighter, but at the same densities as currently achieved using standard cluster amplification.
New compounds and their use as fluorescent labels are provided. The compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications. The labels are advantageous due to their long Stokes shifts. In one embodiment, the compound has the following chemical formula:
C07D 401/06 - Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
C09K 11/07 - Luminescent, e.g. electroluminescent, chemiluminescent, materials containing organic luminescent materials having chemically-interreactive components, e.g. reactive chemiluminescent compositions
C12Q 1/68 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
G01N 33/58 - Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
C09B 23/06 - Methine or polymethine dyes, e.g. cyanine dyes characterised by the methine chain containing an odd number of CH groups three CH groups, e.g. carbocyanines
96.
METHODS, CARRIER ASSEMBLIES, AND SYSTEMS FOR IMAGING SAMPLES FOR BIOLOGICAL OR CHEMICAL ANALYSIS
Method comprising capturing a series of images of overlapping portions of a microarray of features; analyzing light intensities associated with respective features in the images and determining data representations of the images; and combining the data representations of adjacent images based on a comparison of the signal values of the data features of the data representations of the adjacent images, thereby generating a data representation of the microarray; analyzing the data representation of the microarray to determine properties or characteristics of a sample.
97.
METHODS, CARRIER ASSEMBLIES, AND SYSTEMS FOR IMAGING SAMPLES FOR BIOLOGICAL OR CHEMICAL ANALYSIS
Method includes positioning a first carrier assembly on a system stage. The carrier assembly includes a support frame having an inner frame edge that defines a window of the support frame. The first carrier assembly includes a first substrate that is positioned within the window and surrounded by the inner frame edge. The first substrate has a sample thereon. The method includes detecting optical signals from the sample of the first substrate. The method also includes replacing the first carrier assembly on the system stage with a second carrier assembly on the system stage. The second carrier assembly includes the support frame and an adapter plate held by the support frame. The second carrier assembly has a second substrate held by the adapter plate that has a sample thereon. The method also includes detecting optical signals from the sample of the second substrate.
G01N 21/75 - Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
G01N 33/50 - Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
G01N 35/04 - Automatic analysis not limited to methods or materials provided for in any single one of groups ; Handling materials therefor using a plurality of sample containers moved by a conveyor system past one or more treatment or analysis stations - Details of the conveyor system
C40B 40/00 - Libraries per se, e.g. arrays, mixtures
C40B 50/14 - Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
98.
METHODS, CARRIER ASSEMBLIES, AND SYSTEMS FOR IMAGING SAMPLES FOR BIOLOGICAL OR CHEMICAL ANALYSIS
Method includes positioning a first carrier assembly on a system stage. The carrier assembly includes a support frame having an inner frame edge that defines a window of the support frame. The first carrier assembly includes a first substrate that is positioned within the window and surrounded by the inner frame edge. The first substrate has a sample thereon. The method includes detecting optical signals from the sample of the first substrate. The method also includes replacing the first carrier assembly on the system stage with a second carrier assembly on the system stage. The second carrier assembly includes the support frame and an adapter plate held by the support frame. The second carrier assembly has a second substrate held by the adapter plate that has a sample thereon. The method also includes detecting optical signals from the sample of the second substrate.
G01N 21/77 - Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
G01N 21/01 - Arrangements or apparatus for facilitating the optical investigation
G01N 37/00 - INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES - Details not covered by any other group of this subclass
99.
METHODS AND ARRAYS FOR PRODUCING AND SEQUENCING MONOCLONAL CLUSTERS OF NUCLEIC ACID
The invention relates to a microarray comprising: a) a substrate comprising at least one well, a surface surrounding the well and an inner well surface; b) a first layer at least partially covering the inner well surface and comprising at least one first capture primer pair; and c) a second layer covering the first layer and the surface surrounding the well, wherein the first capture primer pair in the first layer is present and functional after the second layer has been provided. The invention also relates to methods for amplifying a nucleic acid.
C12Q 1/6876 - Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
C12P 19/34 - Polynucleotides, e.g. nucleic acids, oligoribonucleotides
C40B 40/06 - Libraries containing nucleotides or polynucleotides, or derivatives thereof
C40B 50/14 - Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
100.
SEQUENCING FROM MULTIPLE PRIMERS TO INCREASE DATA RATE AND DENSITY
The present invention relates to a sequencing method which allows for increased rates of sequencing and an increase in the density of sequencing data. The system may be based on next generation sequencing methods such as sequencing by synthesis (SBS] but uses multiple primers bound at different positions on the same nucleic acid strand.
patents
